Transcriptional regulators of legume-rhizobia symbiosis: Nuclear Factors Ys and GRAS are two for tango

Transcription factors are DNA binding proteins that regulate gene expression. The nitrogen fixing symbiosis established between legume plants and soil bacteria is a complex interaction, in which plants need to integrate signals derived from the symbiont and the surrounding environment to initiate the developmental program of nodule organogenesis and the infection process. Several transcription factors that play critical roles in these processes have been reported in the past decade, including proteins of the GRAS and NF-Y families. Recently, we reported the characterization of a new GRAS domain containing-protein that interacts with a member of the C subunit of the NF-Y family, which plays an important role in nodule development and the progression of bacterial infection during the symbiotic interaction. The connection between transcription factors of these families highlights the significance of multimeric complexes in the fabulous capacity of plants to integrate and respond to multiple environmental stimuli.Instituto de Biotecnologia y Biologia Molecula

A phylogenetically conserved group of nuclear factor-Y transcription factors interact to control nodulation in legumes

The endosymbiotic association between legumes and soil bacteria called rhizobia leads to the formation of a new root-derived organ called the nodule in which differentiated bacteria convert atmospheric nitrogen into a form that can be assimilated by the host plant. Successful root infection by rhizobia and nodule organogenesis require the activation of symbiotic genes that are controlled by a set of transcription factors (TFs). We recently identified Medicago truncatula nuclear factor-YA1 (MtNF-YA1) and MtNF-YA2 as two M. truncatula TFs playing a central role during key steps of the Sinorhizobium meliloti-M. truncatula symbiotic interaction. NF-YA TFs interact with NF-YB and NF-YC subunits to regulate target genes containing the CCAAT box consensus sequence. In this study, using a yeast two-hybrid screen approach, we identified the NF-YB and NF-YC subunits able to interact with MtNF-YA1 and MtNF-YA2. In yeast (Saccharomyces cerevisiae) and in planta, we further demonstrated by both coimmunoprecipitation and bimolecular fluorescence complementation that these NF-YA, -B, and -C subunits interact and form a stable NF-Y heterotrimeric complex. Reverse genetic and chromatin immunoprecipitation-PCR approaches revealed the importance of these newly identified NF-YB and NF-YC subunits for rhizobial symbiosis and binding to the promoter of MtERN1 (for Ethylene Responsive factor required for Nodulation), a direct target gene of MtNF-YA1 and MtNF-YA2. Finally, we verified that a similar trimer is formed in planta by the common bean (Phaseolus vulgaris) NF-Y subunits, revealing the existence of evolutionary conserved NF-Y protein complexes to control nodulation in leguminous plants. This sheds light on the process whereby an ancient heterotrimeric TF mainly controlling cell division in animals has acquired specialized functions in plants.Instituto de Biotecnologia y Biologia Molecula

Transcriptional regulators of legume-rhizobia symbiosis: Nuclear Factors Ys and GRAS are two for tango

Transcription factors are DNA binding proteins that regulate gene expression. The nitrogen fixing symbiosis established between legume plants and soil bacteria is a complex interaction, in which plants need to integrate signals derived from the symbiont and the surrounding environment to initiate the developmental program of nodule organogenesis and the infection process. Several transcription factors that play critical roles in these processes have been reported in the past decade, including proteins of the GRAS and NF-Y families. Recently, we reported the characterization of a new GRAS domain containing-protein that interacts with a member of the C subunit of the NF-Y family, which plays an important role in nodule development and the progression of bacterial infection during the symbiotic interaction. The connection between transcription factors of these families highlights the significance of multimeric complexes in the fabulous capacity of plants to integrate and respond to multiple environmental stimuli.Instituto de Biotecnologia y Biologia Molecula

A phylogenetically conserved group of nuclear factor-Y transcription factors interact to control nodulation in legumes

The endosymbiotic association between legumes and soil bacteria called rhizobia leads to the formation of a new root-derived organ called the nodule in which differentiated bacteria convert atmospheric nitrogen into a form that can be assimilated by the host plant. Successful root infection by rhizobia and nodule organogenesis require the activation of symbiotic genes that are controlled by a set of transcription factors (TFs). We recently identified Medicago truncatula nuclear factor-YA1 (MtNF-YA1) and MtNF-YA2 as two M. truncatula TFs playing a central role during key steps of the Sinorhizobium meliloti-M. truncatula symbiotic interaction. NF-YA TFs interact with NF-YB and NF-YC subunits to regulate target genes containing the CCAAT box consensus sequence. In this study, using a yeast two-hybrid screen approach, we identified the NF-YB and NF-YC subunits able to interact with MtNF-YA1 and MtNF-YA2. In yeast (Saccharomyces cerevisiae) and in planta, we further demonstrated by both coimmunoprecipitation and bimolecular fluorescence complementation that these NF-YA, -B, and -C subunits interact and form a stable NF-Y heterotrimeric complex. Reverse genetic and chromatin immunoprecipitation-PCR approaches revealed the importance of these newly identified NF-YB and NF-YC subunits for rhizobial symbiosis and binding to the promoter of MtERN1 (for Ethylene Responsive factor required for Nodulation), a direct target gene of MtNF-YA1 and MtNF-YA2. Finally, we verified that a similar trimer is formed in planta by the common bean (Phaseolus vulgaris) NF-Y subunits, revealing the existence of evolutionary conserved NF-Y protein complexes to control nodulation in leguminous plants. This sheds light on the process whereby an ancient heterotrimeric TF mainly controlling cell division in animals has acquired specialized functions in plants.Instituto de Biotecnologia y Biologia Molecula

Structure et fonction des complexes protéiques impliquant les facteurs de transcription NF-Y au cours de l'interaction symbiotique entre Medicago truncatula et Sinorhizobium meliloti

Les plantes de la famille des légumineuses présentent la capacité d'interagir avec des bactéries du sol appelées rhizobia. Cette association conduit à la formation d'un nouvel organe racinaire: le nodule au sein duquel, les rhizobia différenciées fixent l'azote atmosphérique en ammonium assimilable par la plante. Ainsi, grâce à cette symbiose, les légumineuses sont indépendantes des engrais azotés ce qui présente un avantage économique et agronomique. Le facteur de transcription MtNF-YA1 a été identifié chez la légumineuse modèle Medicago truncatula comme un régulateur essentiel de la formation des nodules et de la pénétration des rhizobia dans la racine. MtNF-YA1 appartient à la famille des CCAAT-box binding factor qui est conservée chez l'ensemble des eucaryotes. Cette famille de facteurs de transcription présente la particularité d'agir en hétérotrimères en associant les sous-unités NF-YA, NF-YB et NF-YC. Cette forme trimèrique est essentielle au fonctionnement du facteur de transcription et lui permet de lier l'ADN mais aussi d'interagir avec d'autres régulateurs protéiques. Au cours de ce travail, nous avons tout d'abord identifié une protéine très proche de MtNF-YA1 appelée MtNF-YA2 et nous avons démontré que ces deux protéines jouent des rôles partiellement redondant au cours des étapes précoces de la symbiose rhizobienne. La deuxième partie de ce travail a consisté à la caractérisation des protéines interagissant avec ces deux sous-unités NF-YA au cours de la symbiose rhizobienne. Nous avons ainsi décrit des complexes NF-Y trimèriques symbiotiques révélant une certaine conservation chez les légumineuses et identifié deux protéines JAZ interagissant avec les sous-unités NF-Y symbiotiques. Ces études ont permis une meilleure compréhension du rôle des sous-unités NF-Y dans la symbiose rhizobienne ont permis d'identifier de nouveaux acteurs essentiels à la régulation de ce processus et ouvrent de nouvelles perspectives concernant les mécanismes de régulation et le mode d'action de NF-YA1Legumes plants are able to establish a symbiotic interaction with soil-borne bacteria collectively called rhizobia. This symbiosis leads to the formation of a new root organ named nodule in which differentiated rhizobia fix the atmospheric nitrogen into ammonia for the benefit of the host plant. Thanks to this symbiosis, legume plants are independent of nitrogen fertilizers, which confers them an economical and agronomical advantage. The transcription factor MtNF-YA1 has been previously identified in the model legume Medicago truncatula as an essential regulator of nodule formation and rhizobial infection. MtNF-YA1 belongs to the family of CCAAT-box Binding Factor which is conserved in all eukaryotic systems. The NF-Y family is composed by the unrelated NF-YA, NF-YB and NF-YC proteins which assembled into a heterotrimeric complex. This trimeric complex is required for DNA binding and recruitment of further interacting proteins. During this work, we first identified and characterized MtNF-YA2 a protein closely related to MtNF-YA1 and showed that these two proteins share partially redundant functions during the early steps of the rhizobial symbiosis. We then focused on the search of MtNF-YA1 and MtNF-YA2 interacting proteins using non targeted approaches. These studies led to the characterization of several symbiotic heterotrimeric NF-Y complexes that appeared conserved among leguminous plants. We also identified two JAZ proteins interacting with NF-Y subunits casting new light on potential regulation mechanisms of these complexes. This work thus provided new insights concerning the composition and function of NF-YA1-containing protein complexes and opens up new perspectives for the study of their regulation and mode of action

A phylogenetically conserved group of NF-Y transcription factors interact to control nodulation in legumes

The endosymbiotic association between legumes and rhizobia leads to the formation of root nodules in which differentiated bacteria convert atmospheric nitrogen into a form that can be assimilated by the host plant. Successful root infection by rhizobia and nodule organogenesis require the activation of symbiotic genes that are controlled by a set of early transcription factors (TFs). MtNF-YA1 and MtNF-YA2 are two TFs playing partially redundant functions during several steps of the symbiotic interaction between Medicago truncatula and Sinorhizobium meliloti. NF-Y proteins are part of a transcriptional complex composed of three proteins (NF-YA, NF-YB and NF-YC) which bind DNA at CCAAT-boxes, a motif present in most eukaryotic promoters. In plants, each subunit is encoded by small gene families, potentially leading to a multitude of heterotrimeric NF-Y complexes. Here, using yeast two hybrid screenings, we identified the MtNF-YB and MtNF-YC subunits that interact with MtNF-YA1 and A2. Further, we confirmed, both in yeast and in planta, the formation of trimeric NF-Y complexes and showed that these complexes are functional during nodulation using reverse genetic approaches and ChIP-PCR. Finally, as orthologs of the characterized NF-Y subunits also control nodulation in other legumes, we showed in common bean that similar NF-Y trimers could form in planta. Our results suggest that we have identified a group of evolutionary conserved NF-Y proteins that interact to control nodulation in leguminous plants.Fil: Baudin, Maël. Centre National de la Recherche Scientifique; Francia. Centre de Recherche de Nantes. Institut National de la Recherche Agronomique; FranciaFil: Laloum, Tom. Centre National de la Recherche Scientifique; Francia. Centre de Recherche de Nantes. Institut National de la Recherche Agronomique; FranciaFil: Lepage, Agnes. Centre National de la Recherche Scientifique; Francia. Centre de Recherche de Nantes. Institut National de la Recherche Agronomique; FranciaFil: Rípodas, Carolina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Biotecnología y Biología Molecular. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Instituto de Biotecnología y Biología Molecular; ArgentinaFil: Ariel, Federico Damian. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Centre D'etudes de Saclay; FranciaFil: Frances, Lisa. Centre National de la Recherche Scientifique; Francia. Centre de Recherche de Nantes. Institut National de la Recherche Agronomique; FranciaFil: Crespi, Martin. Centre D'etudes de Saclay; FranciaFil: Gamas, Pascal. Centre National de la Recherche Scientifique; FranciaFil: Blanco, Flavio Antonio. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Biotecnología y Biología Molecular. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Instituto de Biotecnología y Biología Molecular; ArgentinaFil: Zanetti, María Eugenia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Biotecnología y Biología Molecular. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Instituto de Biotecnología y Biología Molecular; ArgentinaFil: de Carvalho-Niebel, Fernanda. Centre National de la Recherche Scientifique; Francia. Centre de Recherche de Nantes. Institut National de la Recherche Agronomique; FranciaFil: Niebel, Andreas. Centre National de la Recherche Scientifique; Francia. Centre de Recherche de Nantes. Institut National de la Recherche Agronomique; Franci

Genome‐wide identification of fitness determinants in the Xanthomonas campestris bacterial pathogen during early stages of plant infection

Plant diseases are an important threat to food production. While major pathogenicity determinants required for disease have been extensively studied, less is known on how pathogens thrive during host colonization, especially at early infection stages. Here, we used randomly barcoded-transposon insertion site sequencing (RB-TnSeq) to perform a genome-wide screen and identify key bacterial fitness determinants of the vascular pathogen Xanthomonas campestris pv campestris (Xcc) during infection of the cauliflower host plant (Brassica oleracea). This high-throughput analysis was conducted in hydathodes, the natural entry site of Xcc, in xylem sap and in synthetic media. Xcc did not face a strong bottleneck during hydathode infection. In total, 181 genes important for fitness were identified in plant-associated environments with functional enrichment in genes involved in metabolism but only few genes previously known to be involved in virulence. The biological relevance of 12 genes was independently confirmed by phenotyping single mutants. Notably, we show that XC_3388, a protein with no known function (DUF1631), plays a key role in the adaptation and virulence of Xcc possibly through c-di-GMP-mediated regulation. This study revealed yet unsuspected social behaviors adopted by Xcc individuals when confined inside hydathodes at early infection stages